Contents lists available at ScienceDirect Bioorganic & Medicinal Chemistry journal homepage: www.elsevier.com/locate/bmc C4-Phenylthio β-lactams: Effect of the chirality of the β-lactam ring on antimicrobial activity Rostislav Kuskovsky a,f , Dina Lloyd a,f , Kriti Arora b,g , Balbina J. Plotkin c , Jacalyn M. Green d , Helena I. Boshoff b , Clifton Barry III b , Jeffrey Deschamps e , Monika I. Konaklieva a, ⁎ a Department of Chemistry, American University, Washington, DC 20016, USA b Tuberculosis Research Section, LCIM, NIAID, NIH, 9000 Rockville Pike, Bethesda, MD 20892, USA c Department of Microbiology and Immunology, Midwestern University, Chicago, IL 60515, USA d Department of Biochemistry, Midwestern University, Chicago, IL 60515, USA e Naval Research Laboratory, Code 6930 4555 Overlook Ave., Washington, DC 20375, USA ABSTRACT C4-phenylthio β-lactams are a new family of antibacterial agents that have activity against two phylogenetically distant bacteria – Mycobacterium tuberculosis (Mtb) and Moraxella catarrhalis (M. cat). These compounds are effective against β-lactamase producing Mtb and M. cat unlike the clinically relevant β-lactam antibiotics. The structure-activity relationship for the C4 phenylthio β-lactams has not yet been completely defined. Earlier efforts in our laboratories established that the C4- phenylthio substituent is essential for antimicrobial activity, while the N1 carbamyl substituent plays a more subtle role. In this present study, we investigated the role that the stereochemistry at C4 plays in these compounds’ antibacterial activity. This was achieved by synthesizing and testing the antimicrobial activity of diastereomerswithachiralcarbamylgroupatN1.OurfindingsindicatethatastrictstereochemistryfortheC4-phenylthioβ-lactamsisnotrequiredtoobtainoptimal anti-Mtb and anti-M. cat activity. Furthermore, the structure–bioactivity profiles more closely relate to the electronic requirement of the phenylthiogroup. In addition, the MICs of Mtb are sensitive to growth medium composition. Select compounds showed activity against non-replicating and multi-drug resistant Mtb. 1. Introduction Despite their apparent structural similarities to the penicillins and other members of the β-lactam family of antibiotics, the C4-phenylthio β-lactams 4 (Scheme 1) represent a novel family of antibacterial com- pounds that selectively inhibit the growth of serine β-lactamase pro- ducing Mycobacterium tuberculosis (Mtb) and Moraxella catarrhalis (M. cat). 1,2 These C4-phenylthio β-lactams lack the ionizable group either in the proximity of, (penicillins, cephalosporins, penems, carbapenems) or on, (monobactams), the lactam nitrogen of the β-lactam ring (Fig.1). Until recently, it was generally accepted that for β-lactams to exert bactericidal activity, they must contain a scaffold, which specifically has an ionizable group at the lactam nitrogen within 3.6 Å of the β- lactamcarbonylcarbon.However,therenowappeartobeexceptionsto this scaffold requirement. N-alkylthiolated β-lactams with only lipo- philic substituents on the lactam ring exhibit inhibitory but not cidal antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA) 3 and Bacillus subtilis. 4 The alkylthio substituent on the lactam nitrogen allows for a transfer of the alkylthio group onto a cellular nucleophile. It is hypothesized that the absence of a hydrophilic ring functionality enables activity of the C4 arylthio β-lactams 4 in the presence of β-lactamases. 5 Thus, these compounds operate as bacter- icidal agents under conditions that render most of the familiar β-lactam antibiotics ineffective. This hypothesis is supported by studies demon- strating that once the ionizable group is deleted from the lactam ni- trogen, novel molecular targets emerge. 6–9 We have established that the arylthio substituent at C4 is required for the antibacterial activity; however, the structural features of the type 4 lactams associated with their biological activity have not been completely defined. 1 These compounds can be inactivated by either replacement of the sulfur atom of the phenylthio group at C4 with an oxygen atom, or by replacement of the aryl group of the arylthio sub- stituent at C4 with an alkylthio group. 2 The presence of a sulfur atom coupled with β-unsaturation at C4 is a requirement for the antibacterial activity of these lactams. Prior studies in our laboratory suggest that the antibacterial properties of phenylthio-β-lactams 4 (Scheme 1) are as- sociated with the presence of halogens, or an electron-withdrawing group (EWG) on the phenyl ring of the phenylthio substituent at C4, 1 https://doi.org/10.1016/j.bmc.2019.115050 Received 6 May 2019; Received in revised form 16 July 2019; Accepted 15 August 2019 ⁎ Corresponding author. E-mail address: mkonak@american.edu (M.I. Konaklieva). f Theseauthorscontributedequallytothiswork.D.Lloydpresentaddress:DepartmentofChemistry,TuftsUniversity,62TalbotAvenue,Medford,MA02155,USA; R. Kuskovsky present address: A.J. Drexel Autism Institute, 3020 Market St, Suite 560, Philadelphia, PA 19104, USA. g K. Arora present address: karora@proteus.com. Bioorganic & Medicinal Chemistry 27 (2019) 115050 Available online 20 August 2019 0968-0896/ © 2019 Elsevier Ltd. All rights reserved. T